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RNA Expression (rna + expression)
Kinds of RNA Expression Terms modified by RNA Expression Selected AbstractsMetabolic Acidosis Stimulates RANKL RNA Expression in Bone Through a Cyclo-oxygenase-Dependent Mechanism,JOURNAL OF BONE AND MINERAL RESEARCH, Issue 7 2003Kevin K Frick Abstract Metabolic acidosis inhibits osteoblastic bone formation and stimulates osteoclastic resorption. To determine whether acidosis alters expression of RNA for the osteoclastic differentiation factor RANKL, mouse calvariae were incubated in neutral or physiologically acidic media. Acidosis resulted in a significant cyclo-oxygenase-dependent increase in RANKL RNA levels, which would be expected to induce the associated increase in bone resorption. Introduction: Metabolic acidosis increases net calcium efflux from bone, initially through physicochemical mechanisms and later through predominantly cell-mediated mechanisms. Acidosis decreases osteoblastic bone formation and increases osteoclastic resorption. The growth and maturation of osteoclasts, derived from hematopoietic precursors in the monocyte/macrophage lineage, are dependent on the interplay of a number of factors. Commitment of pre-osteoclasts to osteoclasts is induced by the interaction of the osteoclastic cell-surface receptor RANK with a ligand expressed by osteoblasts, RANKL. The RANK/RANKL interaction not only initiates a differentiation cascade that culminates in mature bone-resorbing osteoclasts but also increases osteoclastic resorptive capacity and survival. Methods: To test the hypothesis that metabolic acidosis increases expression of RANKL, we cultured neonatal mouse calvariae in acidic (initial medium pH ,7.1 and [HCO3,] ,11 mM) or neutral (initial medium pH ,7.5 and [HCO3,] ,25 mM) medium for 24 and 48 h. We determined the relative expression of RANKL RNA by reverse transcriptase-polymerase chain reaction (RT-PCR) and quantitated the expression by Northern analysis. Results: In this model of metabolic acidosis, there was significantly increased expression of RANKL RNA at both 24 (2-fold) and 48 h (5-fold) compared with respective controls. Net calcium efflux from bone was also increased in acidic medium compared with control medium. At 48 h, net calcium efflux correlated directly with RANKL expression (r = 0.77, n = 15, p < 0.001). Inhibition of prostaglandin synthesis with indomethacin blocked the acid-induced increase in RANKL RNA as well as the increased calcium efflux. Conclusions: Metabolic acidosis induces osteoblastic prostaglandin synthesis, followed by autocrine or paracrine induction of RANKL. This increase in RANKL would be expected to augment osteoclastic bone resorption and help explain the increase in cell-mediated net calcium efflux. [source] ORIGINAL ARTICLE: Identification of Toll-Like Receptors in the Rat (Rattus norvegicus): Messenger RNA Expression in the Male Reproductive Tract Under Conditions of Androgen VariationAMERICAN JOURNAL OF REPRODUCTIVE IMMUNOLOGY, Issue 4 2009Barnali Biswas Problem, Although the majority of Toll-like receptors (TLRs) are reported in many species, some of them are not yet described in the rat. Further, factors that govern Tlr expression in the male reproductive tract have received little attention. We attempt to identify and characterize Tlrs in the rat and determine the expression profile under conditions that affect male reproductive tract gene expression. Method of study, Rat Tlr5, Tlr10, and Tlr11 transcript sequences were submitted to GenBank and in silico characterization carried out using bioinformatics tools. RT-PCR analyses using gene specific primers for rat Tlr1,13 were carried out with RNA isolated from reproductive tract tissues of various experimental groups. Results,Tlr5, Tlr10, and Tlr11 identified in this study share features that are characteristic of the known TLRs. Abundant Tlr expression was observed in the male reproductive tract of adult and developing rats. Further, Tlr expression was also observed in the epididymides of androgen ablated rats. Conclusion,Tlr5, Tlr10, and Tlr11 are ubiquitously expressed in the rat. Tlrs seem to be expressed during male reproductive tract development and under conditions of androgen ablation, suggesting the preparedness of the male reproductive tract to detect an infection under all conditions of androgen status. [source] A hypermorphic mouse Gli3 allele results in a polydactylous limb phenotypeDEVELOPMENTAL DYNAMICS, Issue 3 2007Chengbing Wang Abstract Gli3 protein processing to generate the Gli3 repressor is mediated by proteasome and inhibited by Hedgehog signaling. The Gli3 repressor concentration is graded along the anterior,posterior axis of the developing vertebrate limb due to posteriorly restricted Sonic hedgehog expression. In this study, we created a small deletion at the Gli3 locus (Gli3,68), which causes a half reduction in the Gli3 repressor levels and a slightly increased activity of full-length mutant protein in the limb. Mice homozygous for Gli3,68 develop one to two extra partial digits in the anterior of the limb, while mice carrying one copy of the Gli3,68 allele die soon after birth and display seven digits. These phenotypes are more severe than those found in mice lacking one wild-type Gli3 allele. The expression of dHand, Hoxd12, and Hoxd13 is anteriorly expanded in the limb, even though no up-regulation of Gli1 and Ptc RNA expression is detected. These findings suggest that a decrease in the Gli3 repressor level in combination with an increase in Gli3 full-length activity results in more severe digit patterning abnormalities than those caused by a loss of one wild-type Gli3 allele. Developmental Dynamics 236:769,776, 2007. © 2007 Wiley-Liss, Inc. [source] Regulation of the Neurofibromatosis 2 gene promoter expression during embryonic developmentDEVELOPMENTAL DYNAMICS, Issue 10 2006Elena M. Akhmametyeva Abstract Mutations in the Neurofibromatosis 2 (NF2) gene are associated with predisposition to vestibular schwannomas, spinal schwannomas, meningiomas, and ependymomas. Presently, how NF2 is expressed during embryonic development and in the tissues affected by neurofibromatosis type 2 (NF2) has not been well defined. To examine NF2 expression in vivo, we generated transgenic mice carrying a 2.4-kb NF2 promoter driving ,-galactosidase (,-gal) with a nuclear localization signal. Whole-mount embryo staining revealed that the NF2 promoter directed ,-gal expression as early as embryonic day E5.5. Strong expression was detected at E6.5 in the embryonic ectoderm containing many mitotic cells. ,-gal staining was also found in parts of embryonic endoderm and mesoderm. The ,-gal staining pattern in the embryonic tissues was corroborated by in situ hybridization analysis of endogenous Nf2 RNA expression. Importantly, we observed strong NF2 promoter activity in the developing brain and in sites containing migrating cells including the neural tube closure, branchial arches, dorsal aorta, and paraaortic splanchnopleura. Furthermore, we noted a transient change of NF2 promoter activity during neural crest cell migration. While little ,-gal activity was detected in premigratory neural crest cells at the dorsal ridge region of the neural fold, significant activity was seen in the neural crest cells already migrating away from the dorsal neural tube. In addition, we detected considerable NF2 promoter activity in various NF2-affected tissues such as acoustic ganglion, trigeminal ganglion, spinal ganglia, optic chiasma, the ependymal cell-containing tela choroidea, and the pigmented epithelium of the retina. The NF2 promoter expression pattern during embryogenesis suggests a specific regulation of the NF2 gene during neural crest cell migration and further supports the role of merlin in cell adhesion, motility, and proliferation during development. Developmental Dynamics 235:2771,2785, 2006. © 2006 Wiley-Liss, Inc. [source] Comparison of the antilipolytic effects of an A1 adenosine receptor partial agonist in normal and diabetic ratsDIABETES OBESITY & METABOLISM, Issue 2 2009A. K. Dhalla Introduction and Aims:, Elevated plasma free fatty acid (FFA) concentrations play a role in the pathogenesis of type 2 diabetes (2DM). Antilipolytic agents that reduce FFA concentrations may be potentially useful in the treatment of 2DM. Our previous observation that CVT-3619 lowered plasma FFA and triglyceride concentrations in rats and enhanced insulin sensitivity in rodents with dietary-induced forms of insulin resistance suggested that it might be of use in the treatment of patients with 2DM. The present study was undertaken to compare the antilipolytic effects of CVT-3619 in normal (Sprague Dawley, SD) and Zucker diabetic fatty (ZDF) rats. Results:, ZDF rats had significantly higher fat pad weight, glucose, insulin and FFA concentrations than those of SD rats. EC50 values for forskolin-stimulated FFA release from isolated adipocytes from SD and ZDF rats were 750 and 53 nM, respectively (p < 0.05). Maximal forskolin stimulation of FFA release was significantly (p < 0.01) less in ZDF rats (133 ± 60 ,M) compared with SD rats (332 ± 38 ,M). EC50 values for isoproterenol to increase lipolysis in adipocytes from SD and ZDF rats were 2 and 7 nM respectively. Maximal isoproterenol-stimulated lipolysis was significantly (p < 0.01) lower in adipocytes from ZDF rats (179 ± 23 ,M) compared with SD rats (343 ± 27 ,M). Insulin inhibited lipolysis in adipocytes from SD rats with an IC50 value of 30 pM, whereas adipocytes from ZDF rats were resistant to the antilipolytic actions of insulin. In contrast, IC50 values for CVT-3619 to inhibit the release of FFA from SD and ZDF adipocytes were essentially the same (63 and 123 nM respectively). CVT-3619 inhibited lipolysis more than insulin in both SD (86 vs. 46%, p < 0.001) and ZDF (80 vs. 13%, p < 0.001) adipocytes. In in vivo experiments, CVT-3619 (5 mg/kg, PO) lowered FFA to a similar extent in both groups. Plasma concentrations of CVT-3619 were not different in SD and ZDF rats. There was no significant difference in the messenger RNA expression of the A1 receptors relative to ,-actin expression in adipocytes from SD (0.98 ± 0.2) and ZDF rats (0.99 ± 0.3). Conclusion:, The antilipolytic effects of CVT-3619 appear to be independent of insulin resistance and animal model. [source] An evaluation of the etiology of reduced CYP1A1 messenger RNA expression in the Atlantic tomcod from the Hudson River, New York, USA, using reverse transcriptase polymerase chain reaction analysisENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 5 2001Nirmal K. Roy Abstract Adult Atlantic tomcod, Microgadus tomcod, from the Hudson River, New York State, USA, exhibit reduced inducibility of hepatic cytochrome P4501A1 (CYP1A1) mRNA compared with adult tomcod from the cleaner Miramichi River, New Brunswick, Canada, when treated with coplanar polychlorinated biphenyl (PCB) congeners or 2,3,7,8-tetrachlorodibenzo- p -dioxin. In contrast, little difference in CYP1A1 inducibility is observed between tomcod from these two rivers when treated with polycyclic aromatic hydrocarbons (PAHs). We sought to determine if impaired hepatic CYP1A1 inducibility in Hudson River tomcod results from a multigenerational, genetic adaptation or a single generational, physiological acclimation. Embryos and larvae from controlled experimental crosses of Hudson River and Miramichi River parents were exposed for 24 h to water-borne PCB congener 77 (10 ppm), benzo[a]pyrene (BaP; 10 ppm), or dimethysulfoxide, and CYP1A1 expression was assessed in individual larva using competitive reverse transcriptase polymerase chain reaction (RT-PCR) analysis. The CYP1A1 mRNA was significantly induced in larvae from both populations by BaP (47- and 52-fold) and PCB 77 (9- and 22-fold), although levels of expression were higher in offspring of Miramichi matings. Most important, CYP1A1 mRNA was significantly induced by PCB 77 in larvae from Hudson River parents. Concentrations of dioxin, furan, and PCB congeners were measured in livers and eggs of female tomcod from these two locales to quantify the extent of maternal transfer of contaminants. For both rivers, wet-weight contaminant concentrations were significantly higher (4,7 times) in livers than in eggs of the same females, suggesting that a threshold level of contaminants may have to be reached before CYP1A1 transcription is impaired. We conclude that reduced inducibility of hepatic CYP1A1 mRNA in adult tomcod from the Hudson River is most consistent with single-generational acclimation. [source] Microarray analysis suggests the involvement of proteasomes, lysosomes, and matrix metalloproteinases in the response of motor neurons to root avulsionEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 8 2002Jian Hu Abstract We used microarray analysis of RNA expression from punch samples from ventral horn of spinal cord to identify alterations in gene expression in motor neurons 3 days after proximal spinal root avulsion, a traumatic injury that results in the death of 80% of the motor neurons. This analysis identified the anticipated increases in expression of genes coding for proteins involved in the apoptosis cascades and abortive cell cycle re-entry, as well as decreases in expression of genes coding for proteins related to neuronal functional activity, including groups of genes related to energy metabolism, transporter proteins, ion channels, and receptors. It was also found that cathepsins, metalloproteinases, and proteasome-related protein products were highly up-regulated in motor neurons following axotomy. Each of these products represent pathways that have been implicated in other models of neuronal damage, but which have not previously been described as a response to axotomy. [source] Developmental and activity-dependent genomic occupancy profiles of CREB in monkey area V1GENES, BRAIN AND BEHAVIOR, Issue 2 2009J. Lalonde The mammalian neocortex displays significant plastic rearrangement in response to altered sensory input, especially during early postnatal development. It is believed that cyclic AMP-response element-binding (CREB) plays an important role in orchestrating the molecular events that guide neuroplastic change, although the details of its genomic targets during normal postnatal development or in response to sensory deprivation remain unknown. Here, we performed CREB chromatin immunoprecipitation (ChIP) from monkey area V1 tissue and hybridized enriched DNA fragments to promoter microarrays (ChIP chip analysis). Our goal was to determine and categorize the CREB regulon in monkey area V1 at two distinct developmental stages (peak of critical period vs. adulthood) and after 5 days of monocular enucleation (ME) at both ages. Classification of enriched candidates showed that the majority of isolated promoter loci (n = 795) were common to all four conditions. A particularly interesting group of candidates (n = 192) was specific to samples derived from enucleated infant area V1. Gene ontology analysis of CREB targets during early postnatal development showed a subgroup of genes implicated in cytoskeleton-based structural modification. Analysis of messenger RNA expression (quantitative real-time,polymerase chain reaction) of candidate genes showed striking differences in expression profiles between infant and adult area V1 after ME. Our study represents the first extensive genomic analysis of CREB DNA occupancy in monkey neocortex and provides new insight into the multifaceted transcriptional role of CREB in guiding neuroplastic change. [source] Identification of amplified and expressed genes in breast cancer by comparative hybridization onto microarrays of randomly selected cDNA clonesGENES, CHROMOSOMES AND CANCER, Issue 1 2002Jeremy Clark Microarray analysis using sets of known human genes provides a powerful platform for identifying candidate oncogenes involved in DNA amplification events but suffers from the disadvantage that information can be gained only on genes that have been preselected for inclusion on the array. To address this issue, we have performed comparative genome hybridization (CGH) and expression analyses on microarrays of clones, randomly selected from a cDNA library, prepared from a cancer containing the DNA amplicon under investigation. Application of this approach to the BT474 breast carcinoma cell line, which contains amplicons at 20q13, 17q11,21, and 17q22,23, identified 50 amplified and expressed genes, including genes from these regions previously proposed as candidate oncogenes. When considered together with data from microarray expression profiles and Northern analyses, we were able to propose five genes as new candidate oncogenes where amplification in breast cancer cell lines was consistently associated with higher levels of RNA expression. These included the HB01 histone acetyl transferase gene at 17q22,23 and the TRAP100 gene, which encodes a thyroid hormone receptor-associated protein coactivator, at 17q11,21. The results demonstrate the utility of this microarray-based CGH approach in hunting for candidate oncogenes within DNA amplicons. © 2002 Wiley-Liss, Inc. [source] CCR2 promotes hepatic fibrosis in mice,HEPATOLOGY, Issue 1 2009Ekihiro Seki Chemokines and chemokine receptors contribute to the migration of hepatic stellate cells (HSCs) and Kupffer cells, two key cell types in fibrogenesis. Here, we investigate the role of CCR2, the receptor for monocyte chemoattractant protein (MCP)-1, MCP-2, and MCP-3, in hepatic fibrosis. Hepatic CCR2, MCP-1, MCP-2, and MCP-3 messenger RNA expression was increased after bile duct ligation (BDL). Both Kupffer cells and HSCs, but not hepatocytes, expressed CCR2. BDL- and CCl4 -induced fibrosis was markedly reduced in CCR2,/, mice as assessed through collagen deposition, ,-smooth muscle actin expression, and hepatic hydroxyproline content. We generated CCR2 chimeric mice by the combination of clodronate, irradiation, and bone marrow (BM) transplantation allowing full reconstitution of Kupffer cells, but not HSCs, with BM cells. Chimeric mice containing wild-type BM displayed increased macrophage recruitment, whereas chimeric mice containing CCR2,/, BM showed less macrophage recruitment at 5 days after BDL. Although CCR2 expressed in the BM enhanced macrophage recruitment in early phases of injury, CCR2 expression on resident liver cells including HSCs, but not on the BM, was required for fibrogenic responses in chronic fibrosis models. In vitro experiments demonstrated that HSCs deficient in CCR2,/, or its downstream mediator p47phox,/, did not display extracellular signal-regulated kinase and AKT phosphorylation, chemotaxis, or reactive oxygen species production in response to MCP-1, MCP-2, and MCP-3. Conclusion: Our results indicate that CCR2 promotes HSC chemotaxis and the development of hepatic fibrosis. (HEPATOLOGY 2009.) [source] Allelic imbalances and homozygous deletion on 8p23.2 for stepwise progression of hepatocarcinogenesis,,HEPATOLOGY, Issue 2 2009Yutaka Midorikawa Early hepatocellular carcinoma (eHCC) originates from the hepatocytes of chronic liver disease and develops into classical hepatocellular carcinoma (HCC). To identify sequential genetic changes in multistep hepatocarcinogenesis, we analyzed molecular karyotypes using oligonucleotide genotyping 50K arrays. First, 1q21.3-44 gain and loss of heterozygosity (LOH) on 1p36.21-36.32 and 17p13.1-13.3 were frequently observed in eHCC, but not in chronic liver diseases, suggesting that such chromosomal aberrations are early, possibly causative events in liver cancer. Next, we detected 25 chromosomal loci associated with liver cancer progression in five HCCs with nodule-in-nodule appearance, in which the inner nodule develops within eHCC lesion. Using these chromosomal regions as independent variables, decision tree analysis was applied on 14 early and 25 overt HCCs, and extracted combination of chromosomal gains on 5q11.1-35.3 and 8q11.1-24.3 and LOH on 4q11-34.3 and 8p11.21-23.3 as distinctive attributes, which can classify early and overt HCCs recursively. In these four altered regions identified as late events of hepatocarcinogenesis, two tumors in 32 overt HCCs analyzed in the present study and one in a set of independent samples of 36 overt HCCs in our previous study harbored a homozygous deletion near the CSMD1 locus on 8p23.2. CSMD1 messenger RNA expression was decreased in HCC without 8p23.2 deletion, possibly due to hypermethylation of the CpG islands in its promoter region. Conclusion: 1q gain and 1p and 17p LOH are early molecular events, whereas gains in 5q and 8q and LOH on 4q and 8p only occur in advanced HCC, and inactivation of the putative suppressor gene, CSMD1, may be the key event in progression of liver cancer. (HEPATOLOGY 2009.) [source] Adiponectin protects LPS-induced liver injury through modulation of TNF-, in KK-Ay obese miceHEPATOLOGY, Issue 1 2004Takayuki Masaki Adiponectin, an adipocytokine, has been identified in adipose tissue, and its receptors are widely distributed in many tissues, including the liver. The present study was performed to clarify the role of adiponectin in lipopolysaccharide (LPS)-induced liver injury using KK-Ay obese mice. We analyzed the effects of adiponectin pretreatment on liver injury induced by D -galactosamine/LPS (GalN/LPS) in KK-Ay obese mice. GalN/LPS treatment induced significant increases in aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels in the blood, apoptotic and necrotic changes in hepatocytes, and/or showed a high degree of lethality. The GalN/LPS-induced liver injury was more pronounced in KK-Ay obese mice than in lean controls. Pretreatment with adiponectin ameliorated the GalN/LPS-induced elevation of serum AST and ALT levels and the apoptotic and necrotic changes in hepatocytes, resulting in a reduction in lethality. In addition, pretreatment with adiponectin attenuated the GalN/LPS-induced increases in serum and hepatic tumor necrosis factor , (TNF-,) levels and increased peroxisome proliferator-activated receptor (PPAR) , messenger RNA expression in the liver. Furthermore, abdominal macrophages from KK-Ay obese mice pretreated with adiponectin in vitro exhibited decreased LPS-induced TNF-, production compared with controls. Finally, adiponectin pretreatment also ameliorated TNF-,-induced liver injury. In conclusion, these findings suggest that adiponectin prevents LPS-induced hepatic injury by inhibiting the synthesis and/or release of TNF-, of KK-Ay obese mice. (HEPATOLOGY 2004;40:177,184.) [source] Alcohol potentiates hepatitis C virus replicon expressionHEPATOLOGY, Issue 1 2003Ting Zhang Alcohol consumption accelerates liver damage and diminishes the anti-hepatitis C virus (HCV) effect of interferon alfa (IFN-,) in patients with HCV infection. It is unknown, however, whether alcohol enhances HCV replication and promotes HCV disease progression. The availability of the HCV replicon containing hepatic cells has provided a unique opportunity to investigate the interaction between alcohol and HCV replicon expression. We determined whether alcohol enhances HCV RNA expression in the replicon containing hepatic cells. Alcohol, in a concentration-dependent fashion, significantly increased HCV replicon expression. Alcohol also compromised the anti-HCV effect of IFN-,. Investigation of the mechanism(s) responsible for the alcohol action on HCV replicon indicated that alcohol activated nuclear factor ,B (NF-,B) promoter. Caffeic acid phenethyl ester (CAPE), a specific inhibitor of the activation of NF-,B, abolished alcohol-induced HCV RNA expression. In addition, naltrexone, an opiate receptor antagonist, abrogated the enhancing effect of alcohol on HCV replicon expression. In conclusion, alcohol, probably through the activation of NF-,B and the endogenous opioid system, enhances HCV replicon expression and compromises the anti-HCV effect of IFN-,. Thus, alcohol may play an important role in vivo as a cofactor in HCV disease progression and compromise IFN-,-based therapy against HCV infection. [source] Effect of porto-systemic shunting on NOS expression after extended hepatectomy in ratsHEPATOLOGY RESEARCH, Issue 1 2009Hironori Hayashi Aim:, Several surgical procedures have been developed for reducing portal vein pressure to prevent postoperative liver injury. Nitric oxide synthase expression (NOS) induced by elevation of portal vein pressure is thought to play an important role in liver regeneration, but the details are not well understood. Methods:, Rats in the control group and in the subcutaneous splenic transposition (SST) group underwent 90% partial hepatectomy. Survival and portal vein pressure were analyzed. The serum IL-6 and TNF-, levels were measured by enzyme-linked immunosorbent assay (ELISA). Hepatocyte proliferation and apoptosis 12 hours after hepatectomy were analyzed immunohistochemically. The protein and messenger RNA expression of inducible and endothelial NOS were analyzed using Western blotting and quantitative reverse transcriptase polymerase chain reaction, respectively. Results:, The survival rate of the SST group was significantly higher. Portal vein pressure, TNF-, level and the apoptotic index were significantly lower in the SST group. Twelve hours after surgery, liver inducible NOS (iNOS) protein expression was significantly lower in the SST group. However, protein expression of endothelial NOS was not significantly different between the groups. Conclusion:, Inducible NOS expression after extended hepatectomy is related to the effects of porto-systemic shunting on the splanchnic circulation. Also, iNOS induction and concomitant nitric oxide generation appear to participate in the cytotoxicity of excessive portal pressure after extended hepatectomy. [source] Tumor necrosis factor-, augments lipopolysaccharide-induced suppressor of cytokine signalling 3 (SOCS-3) protein expression by preventing the degradationIMMUNOLOGY, Issue 1 2010Jargalsaikhan Dagvadorj Summary The regulatory role of tumour necrosis factor-, (TNF-,) on the expression of suppressor of cytokine signalling 3 (SOCS-3) in response to lipopolysaccharide (LPS) was examined using peritoneal macrophages from TNF-,-deficient mice. The LPS-induced SOCS-3 expression was markedly augmented in macrophages from wild-type mice whereas such augmentation was not seen in the cells from TNF-,-deficient mice. However, there was no significant difference in the level of SOCS-3 messenger RNA expression between macrophages from wild-type mice and those from TNF-,-deficient mice. The addition of exogenous TNF-, augmented the LPS-induced SOCS-3 expression in macrophages from TNF-,-deficient mice. The pulse chase analysis suggested augmented degradation of LPS-induced SOCS-3 protein in macrophages from TNF-,-deficient mice. Moreover, MG 132, a 26S proteasome inhibitor, sustained the LPS-induced SOCS-3 expression in those cells. The tyrosine phosphorylation of SOCS-3 was definitely induced in LPS-stimulated macrophages from TNF-,-deficient mice but not wild-type mice. A tyrosine phosphatase inhibitor enhanced the tyrosine phosphorylation of SOCS-3 in wild-type mice and accelerated the degradation. Therefore, it was suggested that TNF-, prevented the degradation of SOCS-3 protein via inhibition of the tyrosine phosphorylation in LPS-stimulated macrophages. [source] Interleukin-6 is responsible for aberrant B-cell receptor-mediated regulation of RAG expression in systemic lupus erythematosusIMMUNOLOGY, Issue 3 2007Sophie Hillion Summary Defective regulation of secondary immunoglobulin V(D)J gene rearrangement promotes the production of autoantibodies in systemic lupus erythematosus (SLE). It remains unclear, however, whether the regulation of the recombination-activating genes RAG1 and RAG2 is effective in SLE. RAG1 and RAG2 messenger RNA expression was analysed before and after in vitro activation of sorted CD19+ CD5, B cells with anti-immunoglobulin M antibodies, in 20 SLE patients and 17 healthy controls. The expression of CDK2 and p27Kip1 regulators of the RAG2 protein, were examined. The levels of interleukin-6 (IL-6) and its influence on RAG regulation were also evaluated in vitro. SLE patients had increased frequency of RAG-positive B cells. B-cell receptor (BCR) engagement induced a shift in the frequency of ,- and ,-positive cells, associated with a persistence of RAG messenger RNA and the maintenance of RAG2 protein within the nucleus. While expression of the RAG2-negative regulator CDK2 was normal, the positive regulator p27Kip1 was up-regulated and enhanced by BCR engagement. This effect was the result of the aberrant production of IL-6 by SLE B cells. Furthermore, IL-6 receptor blockade led to a reduction in p27Kip1 expression, and allowed the translocation of RAG2 from the nucleus to the cytoplasm. Our study indicates that aberrant production of IL-6 contributes to the inability of SLE B cells to terminate RAG protein production. Therefore, we hypothesize that because of constitutive IL-6 signalling in association with BCR engagement, SLE B cells would become prone to secondary immunoglobulin gene rearrangements and autoantibody production. [source] cDNA cloning, heat shock regulation and developmental expression of the hsp83 gene in the Mediterranean fruit fly Ceratitis capitataINSECT MOLECULAR BIOLOGY, Issue 6 2006M. A. Theodoraki Abstract This report presents the cDNA cloning, heat shock regulation and developmental expression of the hsp90 gene homologue of the Mediterranean fruit fly Ceratitis capitata (medfly). The isolated cDNA contained the coding region, the 3,UTR and most of the 5,UTR of the medfly hsp90 homologue, which was named Cchsp83. The deduced CcHSP83 polypeptide contained all the highly conserved amino acid segments that characterize the cytosolic members of the HSP90 family. Genomic analysis showed that the Cchsp83 gene is unique and was mapped at the 94C division of the sixth polytene chromosome. The size of the Cchsp83 mRNA was found to be approximately 2.7 kb. The predicted molecular mass of the CcHSP83 protein was 81.4 kDa, while the apparent molecular weight estimated by SDS-PAGE was approximately 90 kDa. Phylogenetic analysis based on 14 insect HSP90 amino acid sequences was consistent with the known phylogeny at low taxonomic level. The Cchsp83 gene is constitutively expressed in all stages of medfly development and is induced from a low level to several-fold by heat, depending on the developmental stage. Heat shock induction begins at 30 °C, reaching a maximum between 35 and 41 °C. Cchsp83 RNA expression is highly regulated during embryonic development; however, the temporal fluctuations in RNA levels during embryogenesis were not followed by similar fluctuations in the levels of the protein. [source] Elucidation of a protein signature discriminating six common types of adenocarcinomaINTERNATIONAL JOURNAL OF CANCER, Issue 4 2007Gregory C. Bloom Abstract Pathologists are commonly facing the problem of attempting to identify the site of origin of a metastatic cancer when no primary tumor has been identified, yet few markers have been identified to date. Multitumor classifiers based on microarray based RNA expression have recently been described. Here we describe the first approximation of a tumor classifier based entirely on protein expression quantified by two-dimensional gel electrophoresis (2DE). The 2DE was used to analyze the proteomic expression pattern of 77 similarly appearing (using histomorphology) adenocarcinomas encompassing 6 types or sites of origin: ovary, colon, kidney, breast, lung and stomach. Discriminating sets of proteins were identified and used to train an artificial neural network (ANN). A leave-one-out cross validation (LOOCV) method was used to test the ability of the constructed network to predict the single held out sample from each iteration with a maximum predictive accuracy of 87% and an average predictive accuracy of 82% over the range of proteins chosen for its construction. These findings demonstrate the use of proteomics to construct a highly accurate ANN-based classifier for the detection of an individual tumor type, as well as distinguishing between 6 common tumor types in an unknown primary diagnosis setting. © 2006 Wiley-Liss, Inc. [source] Cigarette smoke extract affects functional activity of MRP1 in bronchial epithelial cellsJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 5 2007Margaretha van der Deen Abstract Cigarette smoke is the principal risk factor for development of chronic obstructive pulmonary disease (COPD). Multidrug resistance-associated protein 1 (MRP1) is a member of the ATP-binding cassette (ABC) superfamily of transporters, which transport physiologic and toxic substrates across cell membranes. MRP1 is highly expressed in lung epithelium. This study aims to analyze the effect of cigarette smoke extract (CSE) on MRP1 activity. In the human bronchial epithelial cell line 16HBE14o,, MRP1 function was studied flow cytometrically by cellular retention of carboxyfluorescein (CF) after CSE incubation and MRP1 downregulation by RNA interference (siRNA). Cell survival was measured by the MTT assay. Immunocytochemically, it was shown that 16HBE14o, expressed MRP1 and breast cancer resistance protein. Coincubation of CSE IC50 (1.53% ± 0.22%) with MK571 further decreased cell survival 31% (p, = 0.018). CSE increased cellular CF retention dose dependently from 1.7-fold at 5% CSE to 10.3-fold at 40% CSE (both p < 0.05). siRNA reduced MRP1 RNA expression with 49% and increased CF accumulation 67% versus control transfected cells. CSE exposure further increased CF retention 24% (p = 0.031). A linear positive relation between MRP1 function and CSE-modulating effects (r = 0.99, p =0.089) was shown in untransfected, control transfected, and MRP1 downregulated 16HBE14o, cells analogous to blocking effects with MRP1 inhibitor MK571 (r = 0.99, p = 0.034). In conclusion, cigarette smoke extract affects MRP1 activity probably competitively in bronchial epithelial cells. Inhibition of MRP1 in turn results in higher CSE toxicity. We propose that MRP1 may be a protective protein for COPD development. © 2007 Wiley Periodicals, Inc. J Biochem Mol Toxicol 21:243,251, 2007; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/jbt.20187 [source] Use of real-time gene-specific polymerase chain reaction to measure RNA expression of three family members of rat cytochrome P450 4AJOURNAL OF BIOCHEMICAL AND MOLECULAR TOXICOLOGY, Issue 3 2001Kimberly B. Bleicher Abstract Exposure of rats to peroxisome proliferators induces members of the cytochrome P450 4A (CYP4A) family. In rats, the CYP4A family consists of four related genes, CYP4A1, CYP4A2, CYP4A3, and CYP4A8. We are specifically interested in examining CYP4A1, CYP4A2, and CYP4A3, each of which is expressed in a tissue-dependent and sex-dependent manner. While CYP4A1 is sufficiently different from the other two members to enable relatively easy specific quantitation, the close similarity between CYP4A2 and CYP4A3 makes quantitative discrimination difficult. We have combined a fluorescent real-time PCR assay (TaqMan®) with the sequence-specific mismatch amplification mutation assay (MAMA) to allow us to carry out specific quantitation of all three members of this family. The assay is designed such that a single fluorescent TaqMan® probe binds to all three gene products, while specificity is conferred by sequence-specific primers. This specific MAMA technique takes advantage of the ability of Taq polymerase to distinguish between the two cDNAs based on mismatches at the 3, end of a PCR primer. In the 84-base PCR product used for this assay, there is only a single-base difference between CYP4A2 and CYP4A3. Despite this similarity, there is at least a 1000-fold discrimination between the two sequences, using CYP4A2 or CYP4A3 specific standards. Analysis of rat liver RNA from both sexes demonstrates that this discrimination is also achieved in complex RNA mixtures. This technique should be broadly applicable to other areas of research such as allelic discrimination, detecting mutational hotspots in tumors, and discrimination among closely related members of other gene families. © 2001 John Wiley & Sons, Inc. J Biochem Mol Toxicol 15:133,142, 2001 [source] Role of atypical protein kinase C isozymes and NF-,B in IL-1,-induced expression of cyclooxygenase-2 in human myometrial smooth muscle cellsJOURNAL OF CELLULAR PHYSIOLOGY, Issue 3 2007Sara V. Duggan Increased myometrial expression of cyclooxygenase-2 (Cox-2) at term results from elevated local levels of inflammatory cytokines, and its inhibition provides a potential route for intervention in human pre-term labor. We have identified a role for atypical protein kinase C (PKC) isozymes in IL-1,-induced Cox-2 expression in human myometrial smooth muscle cells (HMSMC). The PKC inhibitor GF109203X (10 µM) inhibited IL-1,-induced Cox-2 protein and RNA expression, which were also reduced by MAPK and nuclear factor ,B (NF-,B) inhibitors. GF109203X did not affect MAPK activities, and neither did it replicate the effect of p38 MAPK inhibition on Cox-2 mRNA stability, suggesting that PKC operates through an independent mechanism. The effect of GF109203X remained intact after depletion of conventional and novel PKC isozymes by phorbol ester pre-treatment. In contrast LY379196 (10 µM), which at micromolar concentrations inhibits all but atypical PKCs, did not affect Cox-2 expression. A peptide corresponding to the pseudosubstrate sequence of atypical PKCs blocked Cox-2 protein expression, whereas the sequence from conventional PKCs was ineffective. GF109203X did not affect NF-,B binding to nuclear proteins, but strongly reduced NF-,B-dependent transcription in luciferase reporter assays. Our findings indicate that IL-1,-induced Cox-2 expression in HMSMC in culture requires p38-MAPK-mediated mRNA stabilization and an independent activation of Cox-2 transcription which is dependent on the action of atypical PKCs, probably through direct stimulation of the transactivating activity of NF-,B. J. Cell. Physiol. 210: 637,643, 2007. © 2006 Wiley-Liss, Inc. [source] Viral vectors carrying NR1 sequences injected into rat hippocampus interfered with learning and memoryJOURNAL OF NEUROCHEMISTRY, Issue 2002V. Cheli NMDA receptors are relevant to learning and memory as has been shown both by pharmacological and genetic manipulations. Gene knockouts are useful for investigating in vivo functions, but genetic deletions unrestricted in time or region, may lead to developmental defects or death. The challenge is to control expression with temporal and spatial restrictions in the brain. Viral vectors derived from herpes type-1 neurotropic virus are interesting candidates for it. To regulate gene expression of the NMDA receptor NR1 subunit, vectors carrying either sense NR1(+) or antisense NR1(,) sequences and that of the green fluorescent protein (GFP), were constructed. The protein or RNA expression were corroborated in cell culture by GFP autofluorescence, Western blots, immunofluorescence and RT-PCR, and in rat brain, by Western blots and GFP autofluorescence. The vectors were injected into the dorsal hippocampus of adult male Wistar rats. After 6 days each rat was trained and evaluated for both habituation to an open field and inhibitory avoidance to a foot-shock. The rats injected with GFP-NR1(+) vectors showed habituation and learned the inhibitory avoidance, like sham operated rats; while animals injected with GFP-NR1(,) vectors did not. The vectors were useful to modify endogenous gene expression at a defined period, in restricted regions, leading to investigate in vivo functions. NR1 subunit in the hippocampus is involved in mechanisms leading to habituation and to avoidance behaviour, since even a slight change in the availability of NR1 interfered with them. [source] Molecular characterization and expression of maternally expressed gene 3 (Meg3/Gtl2) RNA in the mouse inner earJOURNAL OF NEUROSCIENCE RESEARCH, Issue 2 2006Shehnaaz S.M. Manji Abstract The pathways responsible for sound perception in the cochlea involve the coordinated and regulated expression of hundreds of genes. By using microarray analysis, we identified several transcripts enriched in the inner ear, including the maternally expressed gene 3 (Meg3/Gtl2), an imprinted noncoding RNA. Real-time PCR analysis demonstrated that Meg3/Gtl2 was highly expressed in the cochlea, brain, and eye. Molecular studies revealed the presence of several Meg3/Gtl2 RNA splice variants in the mouse cochlea, brain, and eye. In situ hybridizations showed intense Meg3/Gtl2 RNA staining in the nuclei of type I spiral ganglion cells and in cerebellum near the dorsal vestibular region of the cochlea. In embryonic mouse head sections, Meg3/Gtl2 RNA expression was observed in the otocyst, brain, eye, cartilage, connective tissue, and muscle. Meg3/Gtl2 RNA expression increased in the developing otocyst and localized to the spiral ganglion, stria vascularis, Reissner's membrane, and greater epithelial ridge (GER) in the cochlear duct. RT-PCR analysis performed on cell lines derived from the organ of Corti, representing neural, supporting, and hair cells, showed significantly elevated levels of Meg3/Gtl2 expression in differentiated neural cells. We propose that Meg3/Gtl2 RNA functions as a noncoding regulatory RNA in the inner ear and that it plays a role in pattern specification and differentiation of cells during otocyst development, as well as in the maintenance of a number of terminally differentiated cochlear cell types. © 2005 Wiley-Liss, Inc. [source] RNA from brush oral cytology to measure squamous cell carcinoma gene expressionJOURNAL OF ORAL PATHOLOGY & MEDICINE, Issue 2 2008Joel L. Schwartz Background:, RNA expression analysis of oral keratinocytes can be used to detect early stages of disease such as oral cancer or to monitor on-going treatment responses of the same or other oral diseases. A limitation is the inability to obtain high quality RNA from oral tissue without using biopsies. While oral cytology cell samples can be obtained from patients in a minimally invasive manner they have not been validated for quantitative analysis of RNA expression. Methods:, As a starting point in the analysis of tumor markers in oral squamous cell carcinoma (OSCC), we examined RNA in brush cytology samples from hamsters treated with dibenzo[a,l]pyrene to induce oral carcinoma. Three separate samples from each animal were assessed for expression of candidate marker genes and control genes measured with real-time RT-PCR. Results:, Brush oral cytology samples from normal mucosa were shown to consist almost exclusively of epithelial cells. Remarkably, ß-2 microglobulin and cytochrome p450, 1B1 (CYP1B1) RNA showed potential utility as markers of OSCC in samples obtained in this rapid and non-surgical manner. Conclusion:, Brush oral cytology may prove useful as a source of RNA for gene expression analysis during the progression of diseases of the oral epithelium such as OSCC. [source] Polymethylmethacrylate particles impair osteoprogenitor viability and expression of osteogenic transcription factors Runx2, osterix, and Dlx5JOURNAL OF ORTHOPAEDIC RESEARCH, Issue 5 2010Richard Chiu Abstract Polymethylmethacrylate (PMMA) particles have been shown to inhibit the differentiation of osteoprogenitor cells, but the mechanism of this inhibitory effect has not been investigated. We hypothesize that the inhibitory effects of PMMA particles involve impairment of osteoprogenitor viability and direct inhibition of transcription factors that regulate osteogenesis. We challenged MC3T3-E1 osteoprogenitors with PMMA particles and examined the effects of these materials on osteoprogenitor viability and expression of transcription factors Runx2, osterix, Dlx5, and Msx2. MC3T3-E1 cells treated with PMMA particles over a 72-h period showed a significant reduction in cell viability and proliferation as indicated by a dose- and time-dependent increase in supernatant levels of lactate dehydrogenase, an intracellular enzyme released from dead cells, a dose-dependent decrease in cell number and BrdU uptake, and the presence of large numbers of positively labeled Annexin V-stained cells. The absence of apoptotic cells on TUNEL assay indicated that cell death occurred by necrosis, not apoptosis. MC3T3-E1 cells challenged with PMMA particles during the first 6 days of differentiation in osteogenic medium showed a significant dose-dependent decrease in the RNA expression of Runx2, osterix, and Dlx5 on all days of measurement, while the RNA expression of Msx2, an antagonist of Dlx5-induced osteogenesis, remained relatively unaffected. These results indicate that PMMA particles impair osteoprogenitor viability and inhibit the expression of transcription factors that promote osteoprogenitor differentiation. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 28:571,577, 2010 [source] Young, adult, and old rats have similar changes in mRNA expression of many skeletal genes after fracture despite delayed healing with ageJOURNAL OF ORTHOPAEDIC RESEARCH, Issue 10 2006Ralph A. Meyer Jr. Abstract Genes active in fracture healing are not well understood. Because age slows skeletal repair, the change in gene expression between animals of differing ages may illuminate novel pathways important to this healing response. To explore this, 6-, 26-, and 52-week-old female Sprague-Dawley rats were subjected to mid-diaphyseal femoral fracture with intramedullary fixation. The fracture callus was collected at 0, 0.4 (3 days), 1, 2, 4, or 6 weeks after fracture. RNA was extracted and pooled between two animals for each sample. Three samples were done for each time point for each age for a total of 54 Affymetrix U34A GeneChip microarrays. Of the 8700 genes on each array, 3300 were scored as present. Almost all of these genes were affected by femoral fracture with either upregulation or downregulation in the 6 weeks after fracture. Upregulated genes included markers for matrix genes for both cartilage and bone, osteoblasts, osteocytes, osteoclasts, fibroblasts, and mast cells. Downregulated genes included genes related to blood cell synthesis. Nearly all genes presently associated with bone metabolism showed the same response to fracture healing regardless of the age of the animal. In conclusion, skeletal fracture led to similar changes in RNA expression for most skeletal genes despite the delay in the formation of bone to bridge the fracture gap in old rats. Defects in the healing of skeletal trauma in older rats may lie in systems not normally studied by skeletal biologists. © 2006 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 24:1933,1944, 2006 [source] Melatonin interactions with blood pressure and vascular function during l -NAME-induced hypertensionJOURNAL OF PINEAL RESEARCH, Issue 2 2010Ludovit Paulis Abstract:, The mechanisms responsible for the antihypertensive effect of melatonin are not completely understood. To elucidate the possible role of the nitric oxide (NO) pathway in the hemodynamic actions of melatonin, the effects of this indolamine on vascular function during hypertension induced by the NO-synthase (NOS) inhibitor, N, -nitro- l -arginine-methyl ester (l -NAME) were investigated. Four groups of male adult Wistar rats were employed: control, L-NAME (40 mg/kg), melatonin (10 mg/kg) and l -NAME + melatonin for 5 wks. Systolic and diastolic blood pressure were measured invasively in the carotid artery. Conjugated dienes concentration (an oxidative load marker), NOS RNA expression and its activity and RNA expression of cyclooxygenase-(COX)-1 and COX-2 were determined in the aorta. Acetylcholine-induced responses and their NO-mediated component were evaluated in femoral and mesenteric artery. Moreover, endothelium-derived constricting factor (EDCF)-dependent vasoconstriction and inner diameter were determined in the femoral artery. Chronic l -NAME treatment induced hypertension, elevated the oxidative load and inhibited NOS activity. Moreover, impaired NO-dependent relaxation, augmented EDCF-constriction, increased COX-2 expression and reduced arterial inner diameter were observed. Melatonin added to l -NAME treatment completely prevented elevation of the oxidative load in the aorta. However, melatonin was not able to prevent NOS activity decline, elevation of COX-2 expression or the impairment of vascular responses (except moderate improvement in relaxation of small mesenteric arteries) and it exerted only slight antihypertensive effect. In conclusion, in addition to the reduction of the oxidative load, the restoration of the NO pathway seems to play an important role in the antihypertensive effect of melatonin. [source] Generation of hepatocytes from cultured mouse embryonic stem cellsLIVER TRANSPLANTATION, Issue 10 2003Xiao Ling Kuai Embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass of fertilized blastocysts in vitro. ES cells can be induced to undergo differentiation into potentially all cell types. The aim of this study is to examine the differentiating potential of mouse ES cells into hepatocytes in the presence of retinoic acid (RA), hepatocyte growth factor (HGF), and ,-nerve growth factor (,-NGF). RA, HGF, and ,-NGF were added to the cell culture. Hepatocyte induction was confirmed morphologically, as well as biochemically, through immunohistochemical assays of ,1 -antitrypsin (,1 -AT) and alfafetaprotein (AFP) expression and reverse-transcriptase polymerase chain reaction tests for the presence of albumin, transthyretin, glucose 6 phosphates, hepatic nuclear factor 4, and SAPK/ERK kinase-1 (SEK1) messenger RNA, produced only by functioning hepatocytes. Fifteen days after the addition of HGF and ,-NGF to the cell culture, many epithelioid cells were noticed. ,1 -AT, AFP, albumin, transthyretin, glucose 6 phosphates, hepatic nuclear factor 4, and SEK1 messenger RNA expression also was detected, indicating successful ES cell differentiation into functioning hepatocytes. However, in the presence of RA alone, only transthyretin messenger RNA was positive, whereas no other expression pertaining to functioning hepatocytes could be detected. In the presence of HGF and ,-NGF, mouse ES cells can differentiate into functioning hepatocytes, whereas RA function is limited. [source] Expression of cyclooxygenase-2 in chronic hepatitis B and the effects of anti-viral therapyALIMENTARY PHARMACOLOGY & THERAPEUTICS, Issue 2 2002A. S. L. Cheng Backgound: Cyclooxygenase-2 may play a role in the development of hepatocellular carcinoma, but the relationship between cyclooxygenase-2 and chronic hepatitis B is unknown. Aim: To investigate the expression and cellular localization of cyclooxygenase-2 in chronic hepatitis B patients and the effects of anti-viral therapy. Methods: Using immunohistochemistry, in situ hybridization, Western blot and reverse transcription polymerase chain reaction, protein and messenger RNA expression and cellular localization of cyclooxygenase-2 in 35 chronic hepatitis B patients were assessed. Fourteen histologically normal and non-viral-infected livers were used as controls. The cyclooxygenase-2 immunoreactivities of paired liver biopsies from 12 patients receiving anti-viral therapy were compared. Results: Immunohistochemistry and in situ hybridization revealed that cyclooxygenase-2 expression was confined to hepatocytes. Patients with chronic hepatitis B had significantly higher cyclooxygenase-2 expression compared with controls. The cyclooxygenase-2 expression of hepatitis B e antigen-positive and -negative chronic hepatitis B patients was not significantly different, although the necro-inflammatory activity of the latter group was significantly lower. Over-expression of cyclooxygenase-2 in patients with chronic hepatitis B was further confirmed by Western blot and reverse transcription polymerase chain reaction. Twelve hepatitis B e antigen-positive chronic hepatitis B patients received anti-viral therapy: lamivudine in seven and interferon in five. Despite hepatitis B e antigen seroconversion, disappearance of hepatitis B virus DNA in serum, normalization of liver enzymes and a significant reduction in necro-inflammatory activity in all 12 patients, no significant change in cyclooxygenase-2 expression was found. Conclusions: Chronic hepatitis B is associated with elevated cyclooxygenase-2 levels in hepatocytes, and the over-expression of this enzyme does not reflect inflammatory activity. Up-regulation of cyclooxygenase-2 persists after successful anti-viral therapy. [source] A proteomic analysis of penicillin resistance in Streptococcus pneumoniae reveals a novel role for PstS, a subunit of the phosphate ABC transporterMOLECULAR MICROBIOLOGY, Issue 5 2005Hafid Soualhine Summary Resistance to penicillin is widespread in the Gram-positive bacterium Streptococcus pneumoniae, and while several mutations are known to be implicated in resistance other mechanisms are likely to occur. We used a proteomic screen of two independent mutants in which resistance was selected in vitro. We found a number of differentially expressed proteins including PstS, a subunit of the phosphate ABC transporter of S. pneumoniae. This protein was increased in both mutants, a phenotype correlated to increased RNA expression of the entire phosphate ABC transporter operon. Inactivation of the pstS gene led to increased susceptibility to penicillin in the wild-type strain. To further link the expression of the ABC phosphate transporter with penicillin resistance, we looked at pstS mRNA levels in 12 independent clinical isolates sensitive and resistant to penicillin and found an excellent correlation between resistance and increased expression of pstS. Inactivation of pstS in one of the clinical isolates significantly reduced penicillin resistance. Global approaches are ideally suited for the discovery of novel factors in the biology of resistance. [source] | |||||||||||||||||||||||||||||||